Inorganic pyrophosphatase: We are currently investigating the active site structure of inorganic pyrophosphatase. Chemical modification studies have provided evidence for the involvement of only arginine and tyrosine residues at the active site. Phenyl glyoxal inactivates the enzyme by modification of one or at most two arginine residues and we are currently tyring to isolate the corresponding tyrptic peptide(s). Work along similar lines is being pursued on localizing the site of modification with diazonium (1H) tetrazole, which also inactivates the enzyme. Magnetic resonance and kinetic experiments are also being used to define the enzymatic mechanism of action. Additionally, we are also studying model reactions for metal ion catalysis of phosphoryl transfer. Adenylate affinity labels: We have synthesized and characterized three photolabile derivatives of 3', 5' cyclic AMP and used them to investigate the 3', 5' cyclic AMP binding sites in rabbit muscle phosphofructokinase and in human erythrocyte ghosts. These compounds are now being studied with number of other cAMP receptors. We have extended our approach to synthesize photolabile derivatives of 5' AMP and of puromycin. The 5' AMP derivatives have been shown to incorporate into phosphorylase b and studies of their effects on the allosteric properties of this enzyme are underway. One puromycin derivative incorporates into E. coli ribosomes, and the details of this process are under investigation. BIBLIOGRAPHIC REFERENCES: B.S. Cooperman, E.N. Jaynes, Jr., D. J. Brunswick and M.A. Luddy, "On the Photoincorporation of Puromycin and N-(ethyl-2-diazomalonyl)puromycin into E. coli Ribosomes," Proc. Nat. Acad. Sci. USA., 72, 2974 (1975).